The stability of an organism's genome is governed by complex mechanisms that involve 1) the repair of DNA damage, 2) the regulation of progression through and transition between phases of the cell cycle and 3) the ability to reorganize chromatin structure. Each of these mechanisms is affected by the nature of the DNA damage and the signal transduction stimuli that alter transcriptional and post-translational activities. Defects in key components within these pathways can manifest in a variety of human disease, such as xeroderma pigmentosum with defects in NER repair and lesion bypass, ataxia telangiectasia, with defects in cell cycle control and intracellular signaling, Fanconi anemia with defective chromatin remodeling and replication restart and colon cancer with defects in mismatch or base excision repair. Thus, integrated responses to maintain of genomic stability are critical for long-term survival and organismal fitness. The focus of our investigations is on bis-electrophiles, a class of environmental and endogenous chemicals that upon exposure to DNA, form a complex mixture of lesions that can contribute to the initiation of cancer and premature aging. In the proposed studies, a series of hypotheses will be tested that will accomplish the following: 1) determine the mutagenic and cytotoxic consequences of replication of DNAs containing site-specific base lesions and the role that DNA repair plays in the modulation of the mutagenic potential of these lesions;2) ascertain key genetic components of and the biochemical basis for a pathway to repair of interstrand crosslinks that occurs with high fidelity in the absence of homologous recombination; and 3) ascertain the extent to which the structure of specific DNA lesions that exist in a dynamic equilibrium between various species (monofunctional adduct, intra- and interstrand DNA crosslinks and DNA-protein crosslinks) affect nucleic acid transactions such as repair and replication. These investigations have direct application to human health since exposure to these compounds, whether from sources produced internally or as an environmental pollutant, contribute to cancer and premature aging. Investigations on the identity of the mutagenic and cytotoxic DNA lesions responsible for these diseases and their mechanisms of repair can ultimately lead to improved rational therapeutic designs.